We hypothesize that amyloid Beta-protein (Abeta) assembly is a seminal neuropathogenetic process in Alzheimer's disease (AD) and in cerebral amyloid angiopathy (CAA). If so, controlling Abeta assembly could be of therapeutic value. Over the last decade, we have worked to elucidate the pathways of Abeta self-association in vitro, to identify assembly intermediates, and to evaluate the neurotoxic activities of these structures. This work led to the discovery of the amyloid protofibril, later found to be neurotoxic in vitro and in vivo, and to be linked to an "Arctic" form of AD. Recently, novel chemical cross-linking studies have revealed the existence of smaller oligomeric structures (paranuclei), which are formed rapidly by Abeta (1-42) but not by Abeta (1-40). The strong association of Abeta (1-42) with AD thus may result from Abeta (1-42)-specific assembly events occurring at the earliest stage of self-association, oligomerization. We also have described a novel helix-rich oligomeric assembly intermediate. We predicted that an Asp23->Asn amino acid replacement would affect the rate of formation of this intermediate and of fibrils. Interestingly, the importance of this site has been proven in humans through the discovery of an Iowa kindred suffering from an early onset form of CAA caused by this exact substitution. In this proposal, we will examine the thermodynamics and structural biology of early Abeta assembly reactions in order to understand the fundamental factors controlling these reactions and to characterize the structures formed. In concurrent experiments, we will examine the neurotoxic activities of the Abeta assemblies to establish which may be of most relevance pathobiologically. Our studies will provide a better understanding of the mechanisms of formation and the biological activities of Abeta assemblies and of general principles of amyloid formation and protein misfolding. Our experimental plan comprises four specific aims. Aim 1. To elucidate the thermodynamics of early Abeta assemblies and Abeta fibril formation. Aim 2. To determine the structural features of early Abeta assemblies. Aim 3. To determine the structure and mechanism of action of oligomeric Abeta fibrillogenesis inhibitors. Aim 4. To determine the biological activity of early intermediates.